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Abstract

Background

Glutathione S-transferases (GSTs) have proved to be involved in the detoxifying several
carcinogens and may play an important role in carcinogenesis of cancer. Previous studies
on the association between Glutathione S-transferase M1 (GSTM1) polymorphism and gastric
cancer (GC) risk reported inconclusive results. To get a precise result, we conducted
this present meta-analysis through pooling all eligible studies.

Methods

A comprehensive databases of Pubmed, Embase, Web of Science, and the Chinese Biomedical
Database (CBM) were searched for case–control studies investigating the association
between GSTM1 null genotype and GC risk. Odds ratios (OR) and 95% confidence intervals
(95% CI) were used to assess this possible association. A χ2-based Q-test was used
to examine the heterogeneity assumption. Begg’s and Egger’s test were used to examine
the potential publication bias. The leave-one-out sensitivity analysis was conducted
to determine whether our assumptions or decisions have a major effect on the results
of present work. Statistical analyses were performed with the software program STATA
12.0.

Virtual Slides

Keywords:

GSTM1; Polymorphism; Gastric cancer; Risk; Meta-analysis

Background

Multiple lines of evidence suggested both cumulative effect of environmental risk
factors and genetic susceptibility of the individual contributed to the development
of the cancers [1]. The gene-environment interaction in carcinogenesis is also well reflected by metabolic
enzymes involved in the inactivation and/or detoxification of environmental carcinogens.
Most of the carcinogens are metabolically inactivated by detoxification enzymes. Therefore,
inherited variations in genes encoding the carcinogen-metabolizing enzymes may alter
enzymatic activity and subsequently the carcinogens activation and/or deactivation
[2]. Individual susceptibility to cancer is likely to be affected by the genotypes of
biotransformation enzymes which represent significant ethnic differences in the frequency
of alleles [3].

Human glutathione S-transferases (GSTs) are phase II metabolizing enzymes that play
a key role in protecting against cancer by detoxifying numerous potentially cytotoxic/genotoxic
compounds [4]. The genes encoding the three major GST isoenzymes, GSTM (mu) 1, GSTT (theta) 1,
and GSTP (pi) 1, widely expressed along the human gastrointestinal tract [5], are highly polymorphic. Among the GST isoforms, glutathione S-transferase M1 (GSTM1)
is of particular interest and important because it possesses a present/null polymorphism
and the null genotype has a complete absence of GSTM1 enzyme activity. It has been
observed that GSTM1 null may affect individual susceptibility to cancer [6]. Up to now, numerous researches about the relationship between the polymorphism of
GSTM1 null genotype and GC susceptibility have been conducted. However, the findings
are controversial due to different reasons including the populations selected and
their ethnicities. A recent meta-analysis of 15 studies suggested no association between
the GSTM1 polymorphism and GC susceptibility was found [7]. When they performed the meta-analysis, the pooled sample size was relatively small
and not enough information was available for more exhaustive subgroup analysis. Since
then, additional several studies with a large sample size about this polymorphism
on GC risk have been reported, which would greatly improve the power of the meta-analysis.
In order to get a more precise result, we conducted this present meta-analysis.

Methods

Search strategy for eligible studies

We conducted a comprehensive search through the Pubmed, Embase, Web of Science, and
Chinese Biomedical Data-base (CBM) databases for studies assessing the association
between GSTM1 null genotype and GC risk. The literature strategy used the following
keywords: (“Glutathione S-transferase M1”, “GSTM1” or “GSTM”) and (“gastric cancer”,
“gastric carcinoma”, “stomach cancer” or “stomach carcinoma”). There was no sample
size and language limitation. We evaluated all associated publications to retrieve
the most eligible literatures. All references cited in the included studies were also
hand-searched and reviewed to identify additional published articles not indexed in
common databases. Of the studies with overlapping data published by the same authors,
only the most recent or complete study was included in this meta-analysis.

Inclusion and exclusion criteria

The inclusion criteria of eligible studies were as following: (1) Evaluate the GSTM1
polymorphism and GC risk; (2) Only the case–control studies were considered; (3) The
paper should clearly describe the diagnoses of GC and the sources of cases and controls;
(4) The controls were gastric cancer-free individuals; (5) Reported the frequencies
of GSTM1 polymorphism in both cases and controls or the odds ratio (OR) and its 95%
confidence interval (95% CI) of the association between GSTM1 null genotype and GC
risk. The exclusion criteria were: (1) none case–control studies; (2) control population
including malignant tumor patients; and (3) duplicated publications.

Data extraction

Relevant data were extracted from all the eligible studies independently by two reviewers,
and disagreements were settled by discussion and the consensus was reached among all
reviewers. The main data extracted from the eligible studies were as following: the
first author, year of publication, ethnicity, genotype method, source of the controls,
total numbers of cases and controls, the genotype frequency of GSTM1 polymorphism.
Different ethnicities were mainly categorized as Caucasians, Asians, Africans, and
Mixed. If a study did not specify the ethnicity or if it was not possible to separate
participants according to such phenotype, the group was termed “mixed”. For studies
including subjects of different ethnic populations, data were collected separately
whenever possible and recognized as an independent study.

Quality assessment

Quality of eligible studies in present meta-analysis was assessed using the Newcastle
Ottawa scale (NOS) as recommended by the Cochrane Non-Randomized Studies Methods Working
Group. This instrument was developed to assess the quality of non-randomized studies,
specifically cohort and case–control studies [8]. This instrument was developed to assess the quality of nonrandomized studies, specifically
cohort and case–control studies. Based on the NOS, case–control studies were judged
based on three broad perspectives: selection of study groups (1 criterion), comparability
of study groups (4 criteria), and ascertainment of outcome of interest (3 criteria).
Given the variability in quality of observational studies found on our initial literature
search, we considered studies that met 5 or more of the NOS criteria as high quality
(http://www.ohri.ca/programs/clinical_epidemiology/oxford.aspwebcite) [9].

Statistical methods

We examined the association between GSTM1 null genotype and GC risk by calculating
pooled odds ratio (ORs), 95% confidence intervals (95% CI), and the significance of
the pooled OR was determined by the Z-test. To assess the heterogeneity among the
included studies more precisely, both the chi-square based Q statistic test (Cochran’s
Q statistic) to test for heterogeneity and the I2 statistic to quantify the proportion of the total variation due to heterogeneity
[10,11]. If obvious heterogeneity existed among those included studies (P < 0.05), the random-effect
model (DerSimonian and Laird method) was used to pool the results [12]. When there was no obvious heterogeneity existed among those included studies (P
> 0.05), the fixed-effect model (Mantel-Haenszel’s method) was used to pool the results
[13]. Moreover, subgroup analyses were performed to test whether the effect size varied
by the ethnicity and the source of control population. The kinds of ethnicity were
mainly defined as Caucasians, Asians. Publication bias was investigated with the funnel
plot and its asymmetry suggested risk of publication bias. To evaluate the published
bias, we used Begg’s [14] and Egger’s [15] formal statistical test and by visual inspection of the funnel plot. Furthermore,
the leave-one-out sensitivity analysis was conducted to determine whether our assumptions
or decisions have a major effect on the results of the review by omitting each study
[16]. All statistical tests for this meta-analysis were performed with STATA (version
12.0; Stata Corporation, College Station, TX). A P value less than 0.05 was considered
statistically significant, and all the P values were two sided.

Results

Study characteristics

There were 113 relevant abstracts identified by searching the key words, and 41 studies
were firstly excluded after the careful review of the abstracts, leaving 72 studies
for full publication review (Figure 1). Of those 72 studies, 25 studies were excluded (6 for containing overlapping data,
11 for reviews, 3 for without adequate data, and 5 for on GSTT1 polymorphism). Table 1 listed the main characteristics of eligible studies included in this meta-analysis.
There are 47 case–control studies, including 6,678 cases and 12,912 controls met the
selection criteria [2,17-62]. Among the 47 studies, 24 studies are of Caucasians and 23 studies are of Asians.
There are 25 studies of hospital-based controls and the rest are population-based
controls.

Figure 2.Meta-analysis of the association between GSTT1 null genotype and gastric cancer risk.

Figure 3.Subgroup analyses of the association between GSTT1 null genotype and gastric cancer
risk by the ethnicity.

Figure 4.Subgroup analyses of the association between GSTT1 null genotype and gastric cancer
risk according to the source of controls.

Sensitivity analysis

In order to compare the sensitivity of this meta-analysis, we conducted a leave-one-out
sensitivity analysis. A single study involved in this meta-analysis was evaluated
each time to reflect the influence of the individual data set to pooled ORs. The results
pattern was not impacted by single study (Figure 5).

Figure 5.Sensitive analysis of the pooled ORs and 95% CI for the overall analysis, omitting
each dataset in the meta-analysis.

Publication bias

Begg’s funnel plot and Egger’s test were used to assess the publication bias in this
present work. The Funnel plots’ shape did not reveal obvious evidence of asymmetry
(Figure 6), and the P value of Egger’s test was more than 0.05, providing statistical evidence
for the funnel plots’ symmetry.

Discussion

Gastric cancer is one of the most common malignancies in the world which accounts
for 9.7% of total cancer deaths. Multiple factors have been proved contributed to
the development of GC, including environmental, such as, Helicobacter pylori infection,
Tobacco smoking and individual genetic polymorphism [63,64]. Since the first publication in 1991 by Strange et al. [17] reporting the association between the GSTT1 null genotype and the increased risk
of GC, a large number of epidemiological studies concerning the link between GST gene
polymorphisms and GC risk have been conducted. GSTM1 is generally considered as a
protective enzyme because it detoxifies a number of toxic and carcinogenic substances
such as nitrosamines and PAHs including BPDE [65].

As we all known, meta-analysis has great power to give a more credible results in
one field than individual study through analyzing all the published research works
with the same field [66,67]. Previous epidemiological studies have evaluated the association between the GSTM1
polymorphism and GC risk, but with inconclusive results. Therefore, it is necessary
to perform this meta-analysis to identify the association between GSTM1 polymorphism
and GC risk by combining the relevant studies published to date. Detection of gene
genotype in all kinds of cancer not only in GC patient, which can be used for new
therapeutic targets, will modify the current therapeutic approach. After pooling available
data from all included studies, we found that there was significant association between
this polymorphism and GC risk in over the world population. Our data are in line with
those reported by Saadat et al. [68] and Boccia et al. [69] who observed a significantly increased risk of GC. This association can be explained
by the reduced ability to detoxify the reactive intermediates that react with DNA
because of the lack of GSTM1 enzyme activity [70].

It has been well known that cancer occurrence and mortality varied by ethnicity and
geographic location. Piao et al. [71] suggested it was not associated with GC risk in different populations. In present
work, significant association of GSTM1 polymorphism with GC risk was detected in Asian
populations. However, no association was detected in Caucasians, which in line with
previous meta-analysis conducted by Qiu et al. [72]. When stratified by source of controls, significant association between GSTM1 polymorphism
and GC risk was observed among hospital-based studies. Many factors may contribute
to this result, incompleteness of search, and include the potential false diagnoses
(clinic, documentation, statistical methods). Furthermore, the use of typical control
populations is vitally important, especially for the genetic association studies.
The failure to reach a statistical significance in population-based studies implies
that the selection of representative controls may reduce bias of the results.

Some limitations of this study should be acknowledged. Firstly, there was some heterogeneity
in both the meta-analysis of total 48 studies and the subgroup analyses by ethnicity.
The differences from the selection criteria of cases or controls, the adjusted confounding
variables, and the ethnicity result in the heterogeneity. Secondly, most studies in
the meta-analysis were retrospective design which could suffer more risk of bias owing
to the methodological deficiency of retrospective studies. Those there was no obvious
risk of publication bias in the present meta-analysis, the risks of other potential
bias were unable to be excluded. Some misclassification bias was possible because
most studies could not exclude latent gastric cancer cases in the control group. Therefore,
more studies with prospective design and low risk of other bias are needed to provide
a more precise estimate of the association between GSTM1 null genotype and GC risk.
Finally, we could not address gene-gene and gene-environmental interactions in the
association between GSTM1 null genotype and GC risk.

Conclusion

In conclusion, the meta-analysis with all the eligible studies published up to now,
provides a more precise evidence for the significant association between GSTM1 null
genotype and increased risk of GC. In addition, more individual studies with well
design are needed to further assess the possible gene-gene and gene-environmental
interactions in the association between GSTM1 null genotype and GC risk.

Competing interests

The authors declare that they have no competing interests.

Authors’ contributions

XM, BL and YL conceived and designed the experiments. XM and BL analyzed the data.
XM, BL and YL contributed reagents/materials/analysis tools. XM and YL wrote the paper.
Yong Liu revised the paper. All authors read and approved the final manuscript.

Acknowledgements

We thank all the people who give technical support and useful discussion of the paper.